Apoptosis signal-regulating kinase inhibitors

ABSTRACT

The present invention relates to compounds of Formula (I): 
                         
Wherein variables are as defined above. The compounds have apoptosis signal-regulating kinase (“ASK”) inhibitory activity, and are thus useful in the treatment of ASK1-mediated conditions, including autoimmune disorders, inflammatory diseases, cardiovascular diseases, diabetes, diabetic nephropathy, cardio-renal diseases, including kidney disease, fibrotic diseases, respiratory diseases, COPD, idiopathic pulmonary fibrosis, acute lung injury, acute and chronic liver diseases, and neurodegenerative diseases.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.15/357,878, filed Nov. 21, 2016, now U.S. Pat. No. 9,943,521, which is acontinuation of U.S. patent application Ser. No. 14/711,331, filed May13, 2015, now abandoned, which is a continuation of U.S. patentapplication Ser. No. 14/135,812, filed Dec. 20, 2013, now U.S. Pat. No.9,051,313, which application claims the benefit under 35 U.S.C. § 119(e)to U.S. Provisional Application No. 61/740,777, filed Dec. 21, 2012, theentire contents of which are incorporated herein by reference in theirentirety.

FIELD OF THE INVENTION

The present invention relates to novel compounds having enzymeinhibitory activity, and to their use in the treatment of ASK1-mediatedconditions, including autoimmune disorders, inflammatory diseases,including chronic kidney disease, cardiovascular diseases andneurodegenerative diseases. The invention also relates to methods fortheir preparation, and to pharmaceutical compositions containing suchcompounds.

BACKGROUND

Mitogen-activated protein kinase (MAPK) signaling cascades couplediverse extracellular and intracellular queues to appropriate cellularstress responses, including cell growth, differentiation, inflammation,and apoptosis (Kumar, S., Boehm, J., and Lee., J. C. (2003) Nat. Rev.Drug Dis. 2:717-726; Pimienta, G., and Pascual, J. (2007) Cell Cycle, 6:2826-2632). MAPKs exist in three groups, MAP3Ks, MAP2Ks, and MAPKs,which are sequentially activated. MAPK3s directly respond toenvironmental signals and phosphorylate MAP2Ks, which in turnphosphorylate specific MAPKs. MAPKs then mediated the appropriatecellular response by phosphorylating cellular substrates, includingtranscription factors that regulate gene expression.

Apoptosis signal-regulating kinase 1 (ASK1) is a member of themitogen-activated protein kinase kinase kinase (“MAP3K”) family thatactivates the c-Jun N-terminal protein kinase (“JNK”) and p38 MAP kinase(Ichijo, H., Nishida, E., Irie, K., Dijke, P. T., Saitoh, M., Moriguchi,T., Matsumoto, K., Miyazono, K., and Gotoh, Y. (1997) Science, 275,90-94). ASK1 is activated by a variety of stimuli including oxidativestress, reactive oxygen species (ROS), LPS, TNF-α, FasL, ER stress, andincreased intracellular calcium concentrations (Hattori, K., Naguro, I.,Runchel, C., and Ichijo, H. (2009) Cell Comm. Signal. 7:1-10; Takeda,K., Noguchi, T., Naguro, I., and Ichijo, H. (2007) Annu. Rev. Pharmacol.Toxicol. 48: 1-8.27; Nagai, H., Noguchi, T., Takeda, K., and Ichijo, I.(2007) J. Biochem. Mol. Biol. 40:1-6). ASK1 undergoes activation viaautophosphorylation at Thr838 in response to these signals and in turnphosphorylates MAP2Ks, such as MKK3/6 and MKK4/7, which thenphosphorylate and activates p38 and JNK MAPKs, respectively. ASK2 is arelated MAP3K that shares 45% sequence homology with ASK1 (Wang, X. S.,Diener, K., Tan, T-H., and Yao, Z. (1998) Biochem. Biophys. Res. Commun.253, 33-37. Although ASK2 tissue distribution is restricted, in somecell types ASK1 and ASK2 have been reported to interact and functiontogether in a protein complex (Takeda, K., Shimozono, R., Noguchi, T.,Umeda, T., Morimoto, Y., Naguro, I., Tobiume, K., Saitoh, M., Matsuzawa,A., and Ichijo, H. (2007) J. Biol. Chem. 282: 7522-7531; Iriyama, T., etal. (2009) Embo J. 28: 843-853) In non stressed conditions, ASK1 is keptin an inactive state through binding to its repressor Thioredoxin (Trx)(Saitoh, M., Nishitoh, H., Fuji, M., Takeda, K., Tobiume, K., Sawada,Y., Kawabata, M., Miyazono, K., and Ichijo, H. (1998) Embo J.17:2596-2606), and through association with AKT (Zhang, L., Chen, J. andFu, H. (1999) Proc. Natl. Acad. Sci. U.S.A 96:8511-8515).

Phosphorylation of ASK1 protein can lead to apoptosis or other cellularresponses depending on the cell type. ASK1 activation and signaling havebeen reported to play an important role in a broad range of diseasesincluding neurodegenerative, cardiovascular, inflammatory, autoimmunity,and metabolic disorders. In addition, ASK1 has been implicated inmediating organ damage following ischemia and reperfusion of the heart,brain, and kidney (Watanabe et al. (2005) BBRC 333, 562-567; Zhang etal., (2003) Life Sci 74-37-43; Terada et al. (2007) BBRC 364: 1043-49).Emerging evidence suggests that ASK2, either alone or in a complex withASK1, may play important roles in human diseases as well. Therefore,therapeutic agents that function as inhibitors of ASK1 and ASK2signaling complexes have the potential to remedy or improve the lives ofpatients suffering from such conditions.

U.S. Publication No. 2007/0276050 describes methods for identifying ASK1inhibitors useful for preventing and/or treating cardiovascular diseaseand methods for preventing and/or treating cardiovascular disease in ananimal. The methods comprise administering to the animal an ASK1inhibitor and, optionally, a hypertensive compound.

U.S. Publication No. 2007/0167386 reports a drug for at least one ofprevention and treatment of cardiac failure containing a compound thatinhibits a functional expression of ASK1 protein in a cardiomyocyte, anda method for screening the drug.

WO2009027283 discloses triazolopyridine compounds, methods forpreparation thereof and methods for treating autoimmune disorders,inflammatory diseases, cardiovascular diseases and neurodegenerativediseases.

SUMMARY OF THE INVENTION

The present invention provides novel compounds that function as ASK1inhibitors, compositions and methods of using said novel compounds. In afirst aspect, the invention relates to compounds of Formula (I):

wherein:R¹ is C₁-C₃ alkyl or C₃-C₆ cycloalkyl, wherein the alkyl or cycloalkylis optionally substituted with one to three halogen atoms;R² is hydrogen or C₁-C₆ alkyl wherein the alkyl is optionallysubstituted with halo.R³ is hydrogen or C₁-C₃ alkyl;R⁴ is hydrogen or C₁-C₃ alkyl;R⁵ is hydrogen, C₁-C₃ alkyl, OR^(a) or —NHR^(a);R⁶ is hydrogen, C₁-C₃ alkyl, C₁-C₃ haloalkyl, or C₃-C₆ cycloalkylwherein the cycloalkyl is optionally substituted with C₁-C₃ alkyl, C₁-C₃haloalkyl, or 1 or 2 halogen atoms;R^(a) and R^(b) are independently hydrogen, C₁-C₃ alkyl or R^(a) andR^(b) combine with the nitrogen atom to which they are attached to forma four to six member heterocyclic ring optionally containing an oxygenor a nitrogen atom in the ring;or a pharmaceutically acceptable salt or stereoisomer thereof.

In a second aspect, the invention relates to a method of using thecompounds of Formula (I) in the treatment of a disease or condition in apatient, preferably a human patient, which is amenable to treatment byan ASK1 inhibitor. Such diseases include autoimmune disorders,inflammatory diseases, cardiovascular diseases (including diabetes,diabetic nephropathy, and other complications of diabetes), cardio-renaldiseases, including kidney disease, fibrotic diseases, respiratorydiseases (including COPD, idiopathic pulmonary fibrosis (IPF), and acutelung injury), acute and chronic liver diseases, and neurodegenerativediseases.

In a third aspect, the invention relates to pharmaceutical compositioncomprising a therapeutically effective amount of a compound of Formula(I) and at least one pharmaceutically acceptable excipient.

In a fourth embodiment, the invention provides a compound of formula (I)useful for treating chronic kidney disease.

In a fifth embodiment, the invention provides a compound of formula (I)useful for treating kidney fibrosis

In another aspect, the disclosure provides a compound of formula (I) foruse in therapy.

In another aspect, the disclosure relates to the use of a compound offormula (I) for the manufacture of a medicament for treating autoimmunedisorders, inflammatory diseases, cardiovascular diseases (includingdiabetes, diabetic nephropathy, and other complications of diabetes),cardio-renal diseases, including kidney disease, fibrotic diseases,respiratory diseases (including COPD, idiopathic pulmonary fibrosis(IPF), and acute lung injury), acute and chronic liver diseases, andneurodegenerative diseases.

DETAILED DESCRIPTION OF THE INVENTION

Definitions and General Parameters

As used in the present specification, the following words and phrasesare generally intended to have the meanings as set forth below, exceptto the extent that the context in which they are used indicatesotherwise.

The term “alkyl” refers to a monoradical branched or unbranchedsaturated hydrocarbon chain having the indicated number of carbon atoms.This term is exemplified by groups such as methyl, ethyl, n-propyl,iso-propyl and the like.

The term “substituted alkyl” refers to:

-   -   1) an alkyl group as defined above, having 1, 2, or 3        substituents selected from the indicated groups.

The term “alkylene” refers to a diradical of a branched or unbranchedsaturated hydrocarbon chain, typically having from 1 to 6 carbon atoms(e.g. 1, 2, 3, 4, 5 or 6 carbon atoms). This term is exemplified bygroups such as methylene (—CH₂—), ethylene (—CH₂CH₂—), the propyleneisomers (e.g., —CH₂CH₂CH₂— and —CH(CH₃)CH₂—), and the like.

The term “amino” refers to the group —NH₂. The term substituted aminorefers to an NHR^(a) or NR^(a)R^(b) group wherein R^(a) and R^(b) (whichmay be same or different) represent the substituent groups.

The term “cycloalkyl” refers to cyclic alkyl groups of from 3 to 6carbon atoms having a single cyclic ring. Such cycloalkyl groupsinclude, by way of example, single ring structures such as cyclopropyl,cyclobutyl, cyclopentyl, and the like.

The term “halogen” or “halo” refers to fluoro, bromo, chloro, and iodo.

The term “haloalkyl” refers to alkyl of 1-3 (or as indicated) carbonatoms substituted by 1, 2, or 3 halo atoms or as chemically permissible.Examples of haloalkyl groups include trifluoromethyl,1,1,1-trifluropropanyl, 1,1,1-trifluoroisopropanyl.

The term “acyl” denotes a group —C(O)R, in which R is hydrogen,optionally substituted alkyl, optionally substituted cycloalkyl,optionally substituted heterocyclyl, optionally substituted aryl, andoptionally substituted heteroaryl.

The term “acylhalide” refers to the group —C(O)X where X is a halogenatom preferably chloro.

As used herein and unless otherwise defined, the terms “heterocyclyl”,heterocyclic and “heterocycle” are synonymous and refer to a saturatedor partially unsaturated cyclic group having a single ring and havingfrom 1 to 6 carbon atoms and from 1 to 2 hetero atoms selected fromnitrogen, sulfur, and/or oxygen within the ring. Heterocyclic groups asused herein include tetrahydrofuranyl, morpholino, piperidinyl,piperazino, dihydropyridino, and the like. The term “four to sixmembered heterocyclic” as used herein implies a heterocyclic group asdefined above wherein the total number of ring members is four to sixand adjustment is made for the number of carbon atoms based on thenumber of heteroatoms in the ring.

“Optional” or “optionally” means that the subsequently described eventor circumstance may or may not occur, and that the description includesinstances where said event or circumstance occurs and instances in whichit does not.

A compound of a given Formula (e.g. the “compound of Formula (I)”) isintended to encompass the compounds of the invention as disclosed, andthe pharmaceutically acceptable salts, pharmaceutically acceptableesters, hydrates, polymorphs, and prodrugs of such compounds.

The invention also includes compounds of Formula I in which one or morehydrogen atoms attached to a carbon atom is/are replaced by deuterium.Such compounds exhibit increased resistance to metabolism, and are thususeful for increasing the half life of any compound of Formula I whenadministered to a mammal. See, for example, Foster, “Deuterium IsotopeEffects in Studies of Drug Metabolism”, Trends Pharmacol. Sci.5(12):524-527 (1984). Such compounds are synthesized by means well knownin the art, for example by employing starting materials in which one ormore hydrogen atoms have been replaced by deuterium.

“Isomers” are different compounds that have the same molecular formula.

“Stereoisomers” are isomers that differ only in the way the atoms arearranged in space.

“Enantiomers” are a pair of stereoisomers that are non-superimposablemirror images of each other. A 1:1 mixture of a pair of enantiomers is a“racemic” mixture. The term “(±)” is used to designate a racemic mixturewhere appropriate.

“Diastereoisomers” are stereoisomers that have at least two asymmetricatoms, but which are not mirror-images of each other.

Any formula or structure given herein, including Formula I compounds, isalso intended to represent unlabeled forms as well as isotopicallylabeled forms of the compounds.

The term “therapeutically effective amount” refers to an amount that issufficient to effect treatment, as defined below, when administered to amammalian patient, preferably a human patient, in need of suchtreatment. The therapeutically effective amount will vary depending uponthe subject and disease condition being treated, the weight and age ofthe subject, the severity of the disease condition, the manner ofadministration and the like, which can readily be determined by one ofordinary skill in the art.

The term “treatment” or “treating” means any administration of acompound of the invention or practice of the method of the invention forthe purpose of

-   -   (i) protecting against the disease, that is, causing the        clinical symptoms of the disease not to develop;    -   (ii) inhibiting the disease, that is, arresting the development        of clinical symptoms; and/or    -   (iii) relieving the disease, that is, causing the regression of        clinical symptoms.

In many cases, the compounds of the present invention are capable offorming acid and/or base salts by virtue of the presence of amino and/orcarboxyl groups or groups similar thereto.

The term “pharmaceutically acceptable salt” of a given compound refersto salts that retain the biological effectiveness and properties of thegiven compound, and which are not biologically or otherwise undesirable.Pharmaceutically acceptable base addition salts can be prepared frominorganic and organic bases. Salts derived from inorganic bases include,by way of example only, sodium, potassium, lithium, ammonium, calciumand magnesium salts. Salts derived from organic bases include, but arenot limited to, salts of primary, secondary and tertiary amines, such asalkyl amines, dialkyl amines, trialkyl amines, substituted alkyl amines,di(substituted alkyl) amines, tri(substituted alkyl) amines, alkenylamines, dialkenyl amines, trialkenyl amines, substituted alkenyl aminesand the like. Also included are amines where the two or threesubstituents, together with the amino nitrogen, form a heterocyclic orheteroaryl group.

Pharmaceutically acceptable acid addition salts may be prepared frominorganic and organic acids. Salts derived from inorganic acids includehydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid,phosphoric acid, and the like. Salts derived from organic acids includeacetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid,malic acid, malonic acid, succinic acid, maleic acid, fumaric acid,tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid,methanesulfonic acid, ethanesulfonic acid, p-toluene-sulfonic acid,salicylic acid, and the like.

As used herein, “pharmaceutically acceptable carrier” includes any andall solvents, dispersion media, coatings, antibacterial and antifungalagents, isotonic and absorption delaying agents and the like. The use ofsuch media and agents for pharmaceutically active substances is wellknown in the art. Except insofar as any conventional media or agent isincompatible with the active ingredient, its use in the therapeuticcompositions is contemplated. Supplementary active ingredients can alsobe incorporated into the compositions.

“Coronary diseases” or “cardiovascular diseases” refer to diseases ofthe cardiovasculature arising from any one or more than one of, forexample, heart failure (including congestive heart failure, diastolicheart failure and systolic heart failure), acute heart failure,ischemia, recurrent ischemia, myocardial infarction, arrhythmias, angina(including exercise-induced angina, variant angina, stable angina,unstable angina), acute coronary syndrome, diabetes, and intermittentclaudication.

“Intermittent claudication” means the pain associated with peripheralartery disease. “Peripheral artery disease” or PAD is a type ofocclusive peripheral vascular disease (PVD). PAD affects the arteriesoutside the heart and brain. The most common symptom of PAD is a painfulcramping in the hips, thighs, or calves when walking, climbing stairs,or exercising. The pain is called intermittent claudication. Whenlisting the symptom intermittent claudication, it is intended to includeboth PAD and PVD.

Arrhythmia refers to any abnormal heart rate. Bradycardia refers toabnormally slow heart rate whereas tachycardia refers to an abnormallyrapid heart rate. As used herein, the treatment of arrhythmia isintended to include the treatment of supra ventricular tachycardias suchas atrial fibrillation, atrial flutter, AV nodal reentrant tachycardia,atrial tachycardia, and the ventricular tachycardias (VTs), includingidiopathic ventricular tachycardia, ventricular fibrillation,pre-excitation syndrome, and Torsade de Pointes (TdP).

“Pharmaceutically-acceptable” means suitable for use in pharmaceuticalpreparations, generally considered as safe for such use, officiallyapproved by a regulatory agency of a national or state government forsuch use, or being listed in the U. S. Pharmacopoeia or other generallyrecognized pharmacopoeia for use in animals, and more particularly inhumans.

“Prodrug” is a compound that, upon in vivo administration, ismetabolized by one or more steps or processes or otherwise converted tothe biologically, pharmaceutically or therapeutically active form of thecompound. For example, prodrug means a compound that is chemicallydesigned to efficiently liberate the partent drug after overcomingbiological barriers to oral delivery. To produce a prodrug, thepharmaceutically active compound is modified such that the activecompound will be regenerated by metabolic processes. The prodrug may bedesigned to alter the metabolic stability or the transportcharacteristics of a drug, to mask side effects or toxicity, to improvethe flavor of a drug or to alter other characteristics or properties ofa drug. By virtue of knowledge of pharmacodynamic processes and drugmetabolism in vivo, those of skill in this art, once a pharmaceuticallyactive compound is known, can design prodrugs of the compound (see,e.g., Nogrady (1985) Medicinal Chemistry A Biochemical Approach, OxfordUniversity Press, New York, pages 388-392).

“Polymorph” refers to the different crystal forms of a compound,resulting from the possibility of at least two different arrangements ofthe molecules of the compound in the solid state. Polymorphs of a givencompound will be different in crystal structure but identical in liquidor vapor states. Different polymorphic forms of a given substance maydiffer from each other with respect to one or more physical properties,such as solubility and dissociation, true density, crystal shape,compaction behavior, flow properties, and/or solid state stability. Oneof skill in the art is aware of processes for generating polymoprhs ofcrystalline compounds. Thus, polymorphic forms of the compounds of theinvention are within the ambit of the invention.

In another embodiment the present invention provides a compound offormula (I) wherein:

R¹ is C₁-C₃ alkyl or C₃-C₆ cycloalkyl, wherein the alkyl or cycloalkylis optionally substituted with one to three halogen atoms;

R² is hydrogen;

R³ is hydrogen;

R⁴ and R⁵ are both hydrogen;

R⁶ is C₁-C₃ alkyl, or C₃-C₆ cycloalkyl;

R^(a) and R^(b) are independently hydrogen or C₁-C₃ alkyl; or R^(a) andR^(b) combine with the nitrogen atom to which they are attached to forma four to six member heterocyclic group optionally containing an oxygenor a nitrogen atom in the ring; wherein the heterocyclic group isfurther optionally substituted with one or two groups independentlyselected from C₁-C₃ alkyl and hydroxyl;or a pharmaceutically acceptable salt or stereoisomer thereof.

In another embodiment, the present invention provides a compound offormula (I) wherein:

R¹ is cyclopropyl;

R² is hydrogen;

R³ is hydrogen;

R⁴ and R⁵ are both hydrogen;

R⁶ is cyclopropyl;

R^(a) and R^(b) are both C₁-C₃ alkyl or R^(a) and R^(b) combined withthe nitrogen atom to which they are attached to form a four to sixmember heterocyclic group optionally containing an oxygen atom in thering; wherein the heterocyclic group is further optionally substitutedwith one or two groups selected from C₁-C₃ alkyl and hydroxyl;or a pharmaceutically acceptable salt or stereoisomer thereof.

In another embodiment, the present invention provides a compound offormula (I) wherein:

R¹ is C₁-C₃ haloalkyl;

R² is hydrogen;

R³ is hydrogen;

R⁴ and R⁵ are both hydrogen;

R⁶ is cyclopropyl;

R^(a) and R^(b) are both C₁-C₃ alkyl or R^(a) and R^(b) combined withthe nitrogen atom to which they are attached to form a four to sixmember heterocyclic group optionally containing an oxygen atom in thering; wherein the heterocyclic group is further optionally substitutedwith one or two groups selected from C₁-C₃ alkyl and hydroxyl;or a pharmaceutically acceptable salt or stereoisomer thereof.

In another embodiment, the present invention provides a compound offormula (I) wherein:

R¹ is C₁-C₃ alkyl or cyclopropyl, wherein the alkyl group is substitutedwith three fluoro atoms;

R² is hydrogen;

R³ is hydrogen;

R⁴ and R⁵ are both hydrogen;

R⁶ is cyclopropyl;

R^(a) and R^(b) are both C₁-C₃ alkyl or R^(a) and R^(b) combined withthe nitrogen atom to which they are attached to form a four to sixmember heterocyclic group optionally containing an oxygen atom in thering; wherein the heterocyclic group is further optionally substitutedwith one or two groups selected from C₁-C₃ alkyl and hydroxyl;or a pharmaceutically acceptable salt or stereoisomer thereof.

In another aspect, the present invention provides a compound of formula(I) wherein:

R¹ is C₁-C₃ alkyl or cyclopropyl, wherein the alkyl group is substitutedwith three fluoro atoms;

R² is hydrogen;

R³ is hydrogen;

R⁴ and R⁵ are both hydrogen;

R⁶ is cyclopropyl;

R^(a) and R^(b) are both methyl; or R^(a) and R^(b) combine with thenitrogen atom to which they are attached to form a morpholino group oran azetidinyl group wherein the azetidinyl group is dialkylated on onecarbon atom, or substituted on one carbon with hydroxyl and a methylgroup; or a pharmaceutically acceptable salt or stereoisomer thereof.

In another embodiment, the present invention provides a compound offormula (I) wherein:

R¹ is C₁-C₃ alkyl or cyclopropyl, wherein the alkyl group is substitutedwith three fluoro atoms;

R² is hydrogen;

R³ is hydrogen;

R⁴ and R⁵ are both hydrogen;

R⁶ is cyclopropyl;

R^(a) and R^(b) are both C₁-C₃ alkyl or R^(a) and R^(b) combined withthe nitrogen atom to which they are attached to form a four to sixmember heterocyclic group optionally containing an oxygen atom in thering; wherein the heterocyclic group is further optionally substitutedwith one or two groups selected from C₁-C₃ alkyl and hydroxyl;or a pharmaceutically acceptable salt or stereoisomer thereof.

In some embodiment, the present invention provides a compound of formula(I) wherein:

R¹ is C₁-C₃ alkyl or C₁-C₃ cycloalkyl, wherein alkyl or cycloalkyl isoptionally substituted with one to three fluoro atoms;

R² is hydrogen;

R³ is hydrogen;

R⁴ is hydrogen;

R⁵ is hydrogen;

R⁶ is C₁-C₃ alkyl, C₁-C₃ haloalkyl, or C₁-C₃ cycloalkyl, whereincycloalkyl is optionally substituted with one to two fluoro atoms;

R^(a) and R^(b) are combined with the nitrogen atom to which they areattached to form a heterocyclic group selected from the group consistingof azetidinyl, pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl;wherein the heterocyclic group is further optionally substituted withone or two groups selected from C₁-C₃ alkyl and hydroxyl;or a pharmaceutically acceptable salt or stereoisomer thereof.

In some embodiment, the present invention provides a compound of formula(I) wherein:

R¹ is methyl, ethyl, propyl, fluoromethyl, difluormethyl,trifluoromethyl, fluoroethyl, difluoroethyl, trifluoroethyl,fluoropropyl, difluoropropyl, trifluoropropyl, fluorocyclopropyl,difluorocyclopropyl, or cyclopropyl;

R² is hydrogen;

R³ is hydrogen;

R⁴ is hydrogen;

R⁵ is hydrogen;

R⁶ is methyl, ethyl, propyl, fluoromethyl, difluormethyl,trifluoromethyl, fluoroethyl, difluoroethyl, trifluoroethyl,fluoropropyl, difluoropropyl, trifluoropropyl, fluorocyclopropyl,difluorocyclopropyl, or cyclopropyl;

R^(a) and R^(b) are combined with the nitrogen atom to which they areattached to form a heterocyclic group selected from the group consistingof azetidinyl, pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl;wherein the heterocyclic group is further optionally substituted withone or two groups selected from methyl, ethyl, propyl, and hydroxyl;or a pharmaceutically acceptable salt or stereoisomer thereof.

The compounds of the invention include, but are not limited to, thosecompounds named below:

4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-morpholinopicolinamide(Compound 1)

4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-(3-hydroxy-3-methylazetidin-1-yl)picolinamide(Compound 2)

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-morpholino-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 3)

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(3-hydroxy-3-methylazetidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 4)

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(dimethylamino)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 5)

5-(azetidin-1-yl)-4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]pyridine-2-carboxamide(Compound 6)

(S)-5-(azetidin-1-yl)-4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 7)

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(pyrrolidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 8)

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-pyrrolidin-1-ylpyridine-2-carboxamide(Compound 9)

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(3-hydroxypyrrolidin-1-yl)pyridine-2-carboxamide(Compound 10)

4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(3-hydroxypyrrolidin-1-yl)-N-(2-(4-((S)-1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 11)

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-piperidin-1-ylpyridine-2-carboxamide(Compound 12)

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(piperidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 13)

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(4-hydroxy-4-methylpiperidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 14)

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(4-hydroxy-4-methylpiperidin-1-yl)pyridine-2-carboxamide(Compound 15)

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(4-methylpiperazin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 16)

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(4-methylpiperazin-1-yl)pyridine-2-carboxamide(Compound 17)

In one aspect, the present application provides compounds, methods andcompositions that use or include a racemic mixture, a mixture containingan enantiomeric excess (e.e.) of one enantiomer, or a mixture containinga diastereomeric excess (d.e.) of one diastereomer. The compounds offormula (I) may comprise at least 1%, at least 10%, at least 20%, atleast 30%, at least 40%, at least 50%, at least 60%, at least 65%, atleast 70%, at least 75%, at least 80%, at least 85%, at least 90%, atleast 95%, at least 98%, or at least 99% of one stereoisomer that hasthe desired ASK-1 activities. By way of example, all chiral aminestarting materials (e.g. R^(a)R^(b)NH) used in general preparation ofcompounds of formula (I) (e.g. Compounds 3, 4, 5, 7, 8, 11, 13, 14, 16)were used as a mixture of enantiomers. In another example, IntermediateE in the general preparation of the compounds of formula (I) wasprepared and used as a mixture containing predominantly(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoro-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide.

In another aspect, the present application provides methods,compositions that use or include optical isomers, racemates, or othermixtures thereof, of the compounds of formula I or a pharmaceuticallyacceptable salt, prodrug, or solvate thereof. The single enantiomer ordiastereomer, i.e., optically active form, may be obtained by asymmetricsynthesis or by resolution of the racemate. Resolution of racemates maybe accomplished, for example, by known methods such as crystallizationin the presence of a resolving agent, or chromatography, using, forexample a chiral high pressure liquid chromatography (HPLC) column. Forexample, there are Z- and E-forms (or cis- and trans-forms) of thecompounds or a pharmaceutically acceptable salt, prodrug, or solvatethereof with carbon-carbon double bonds.

Nomenclature

Names of compounds of the present invention are provided using thenaming system from ChemBioDraw Ultra 11 or the Molecule to Chemical Namecomponent of Pipeline Pilot version 9.1.0 set to International Union ofPure and Applied Chemistry (IUPAC). Other compounds or radicals may benamed with common names, or systematic or non-systematic names. Thenaming and numbering of the compounds of the invention is illustratedwith a representative compound of Formula (I)

which is named:(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(dimethylamino)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide.In another example, the compound having the below structure of

may be referred to as4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(4-methylpiperazin-1-yl)pyridine-2-carboxamideusing the naming system of the Molecule to Chemical Name component ofPipeline Pilot version 9.1.0 set to IUPAC-style names or4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-(4-methylpiperazin-1-yl)picolinamideusing the name system of ChemBioDraw Ultra.Combination Therapy

Human patients being treated for an acute cardiovascular disease eventby administration of ASK1 inhibitors often exhibit diseases orconditions that benefit from treatment with other therapeutic agents.These diseases or conditions can be of the cardiovascular nature or canbe related to pulmonary disorders, metabolic disorders, gastrointestinaldisorders and the like. Additionally, some coronary patients beingtreated for an acute cardiovascular disease event by administration ofan ASK1 inhibitor exhibit conditions that can benefit from treatmentwith therapeutic agents that are antibiotics, analgesics, and/orantidepressants and anti-anxiety agents.

Cardiovascular related diseases or conditions that can benefit from acombination treatment of ASK1 inhibitors with other therapeutic agentsinclude, without limitation, angina, including stable angina, unstableangina (UA), exercised-induced angina, variant angina, arrhythmias,intermittent claudication, myocardial infarction including non-STEmyocardial infarction (NSTEMI), heart failure including congestive (orchronic) heart failure, acute heart failure, or recurrent ischemia.

Therapeutic agents suitable for treating cardiovascular related diseasesor conditions include anti-anginals, heart failure agents,antithrombotic agents, antiarrhythmic agents, antihypertensive agents,and lipid lowering agents.

The co-administration of ASK1 inhibitors with therapeutic agentssuitable for treating cardiovascular related conditions allowsenhancement in the standard of care therapy the patient is currentlyreceiving.

Anti-anginals include beta-blockers, calcium channel blockers, andnitrates. Beta blockers reduce the heart's need for oxygen by reducingits workload resulting in a decreased heart rate and less vigorous heartcontraction. Examples of beta-blockers include acebutolol, atenolol,betaxolol, bisoprolol/hydrochlorothiazide, bisoprolol, carteolol,esmolol, labetalol, metoprolol, nadolol, propranolol, sotalol(Betapace), and timolol.

Nitrates dilate the arteries and veins thereby increasing coronary bloodflow and decreasing blood pressure. Examples of nitrates includenitroglycerin, nitrate patches, isosorbide dinitrate, andisosorbide-5-mononitrate.

Calcium channel blockers prevent the normal flow of calcium into thecells of the heart and blood vessels causing the blood vessels to relaxthereby increasing the supply of blood and oxygen to the heart. Examplesof calcium channel blockers include amlodipine, bepridil, diltiazem,felodipine, nifedipine, nimodipine (Nimotop), nisoldipine, verapamil,and nicardipine.

Agents used to treat heart failure include diuretics, ACE inhibitors,vasodilators, and cardiac glycosides. Diuretics eliminate excess fluidsin the tissues and circulation thereby relieving many of the symptoms ofheart failure. Examples of diuretics include hydrochlorothiazide,metolazone, furosemide, bumetanide, spironolactone, and eplerenone.

Angiotensin converting enzyme (ACE) inhibitors reduce the workload onthe heart by expanding the blood vessels and decreasing resistance toblood flow. Examples of ACE inhibitors include benazepril, captopril,enalapril, fosinopril, lisinopril, moexipril, perindopril, quinapril,ramipril, and trandolapril.

Vasodilators reduce pressure on the blood vessels by making them relaxand expand. Examples of vasodilators include hydralazine, diazoxide,prazosin, clonidine, and methyldopa. ACE inhibitors, nitrates, potassiumchannel activators, and calcium channel blockers also act asvasodilators.

Cardiac glycosides are compounds that increase the force of the heart'scontractions. These compounds strengthen the pumping capacity of theheart and improve irregular heartbeat activity. Examples of cardiacglycosides include digitalis, digoxin, and digitoxin.

Antithrombotics inhibit the clotting ability of the blood. There arethree main types of antithrombotics—platelet inhibitors, anticoagulants,and thrombolytic agents.

Platelet inhibitors inhibit the clotting activity of platelets, therebyreducing clotting in the arteries. Examples of platelet inhibitorsinclude acetylsalicylic acid, ticlopidine, clopidogrel, dipyridamole,cilostazol, persantine sulfinpyrazone, dipyridamole, indomethacin, andglycoprotein llb/llla inhibitors, such as abciximab, tirofiban, andeptifibatide. Beta blockers and calcium channel blockers also have aplatelet-inhibiting effect.

Anticoagulants prevent blood clots from growing larger and prevent theformation of new clots. Examples of anticoagulants include bivalirudin,warfarin, unfractionated heparin, low molecular weight heparin,danaparoid, lepirudin, and argatroban.

Thrombolytic agents act to break down an existing blood clot. Examplesof thrombolytic agents include streptokinase, urokinase, andtenecteplase, and tissue plasminogen activator.

Antiarrhythmic agents are used to treat disorders of the heart rate andrhythm. Examples of antiarrhythmic agents include amiodarone,procainamide, lidocaine, and propafenone. Cardiac glycosides and betablockers are also used as antiarrhythmic agents.

Antihypertensive agents are used to treat hypertension, a condition inwhich the blood pressure is consistently higher than normal.Hypertension is associated with many aspects of cardiovascular disease,including congestive heart failure, atherosclerosis, and clot formation.

Examples of antihypertensive agents include alpha-1-adrenergicantagonists, such as prazosin (Minipress), doxazosin mesylate (Cardura),prazosin hydrochloride (Minipress), prazosin, polythiazide (Minizide),and terazosin hydrochloride (Hytrin); beta-adrenergic antagonists, suchas propranolol (Inderal), nadolol (Corgard), timolol (Blocadren),metoprolol (Lopressor), and pindolol (Visken); centralalpha-adrenoceptor agonists, such as clonidine hydrochloride (Catapres),clonidine hydrochloride and chlorthalidone (Clorpres, Combipres),guanabenz Acetate (Wytensin), guanfacine hydrochloride (Tenex),methyldopa (Aldomet), methyldopa and chlorothiazide (Aldoclor),methyldopa and hydrochlorothiazide (Aldoril); combinedalpha/beta-adrenergic antagonists, such as labetalol (Normodyne,Trandate), Carvedilol (Coreg); adrenergic neuron blocking agents, suchas guanethidine (Ismelin), reserpine (Serpasil); central nervoussystem-acting antihypertensives, such as clonidine (Catapres),methyldopa (Aldomet), guanabenz (Wytensin); anti-angiotensin II agents;ACE inhibitors, such as perindopril (Aceon) captopril (Capoten),enalapril (Vasotec), lisinopril (Prinivil, Zestril); angiotensin-IIreceptor antagonists, such as Candesartan (Atacand), Eprosartan(Teveten), Irbesartan (Avapro), Losartan (Cozaar), Telmisartan(Micardis), Valsartan (Diovan); calcium channel blockers, such asverapamil (Calan, Isoptin), diltiazem (Cardizem), nifedipine (Adalat,Procardia); diuretics; direct vasodilators, such as nitroprusside(Nipride), diazoxide (Hyperstat IV), hydralazine (Apresoline), minoxidil(Loniten), verapamil; and potassium channel activators, such asaprikalim, bimakalim, cromakalim, emakalim, nicorandil, and pinacidil.

Lipid lowering agents are used to lower the amounts of cholesterol orfatty sugars present in the blood. Examples of lipid lowering agentsinclude bezafibrate (Bezalip), ciprofibrate (Modalim), and statins, suchas atorvastatin (Lipitor), fluvastatin (Lescol), lovastatin (Mevacor,Altocor), mevastatin, pitavastatin (Livalo, Pitava) pravastatin(Lipostat), rosuvastatin (Crestor), and simvastatin (Zocor).

Patients in need of the ASK1 inhibitor often suffers from secondarymedical conditions such as one or more of a metabolic disorder, apulmonary disorder, a peripheral vascular disorder, or agastrointestinal disorder. Such patients can benefit from treatment of acombination therapy comprising administering to the patient thecompounds of the invention in combination with at least one therapeuticagent.

Pulmonary disorder refers to any disease or condition related to thelungs. Examples of pulmonary disorders include, without limitation,asthma, chronic obstructive pulmonary disease (COPD), bronchitis, andemphysema.

Examples of therapeutics agents used to treat pulmonary disordersinclude bronchodilators including beta2 agonists and anticholinergics,corticosteroids, and electrolyte supplements. Specific examples oftherapeutic agents used to treat pulmonary disorders includeepinephrine, terbutaline, albuterol, salmeterol, Serevent, theophylline,ipratropium bromide, tiotropium, methylprednisolone, magnesium, andpotassium.

Examples of metabolic disorders include, without limitation, diabetes,including type I and type II diabetes, metabolic syndrome, dyslipidemia,obesity, glucose intolerance, hypertension, elevated serum cholesterol,and elevated triglycerides.

Examples of therapeutic agents used to treat metabolic disorders includeantihypertensive agents and lipid lowering agents, as described in thesection “Cardiovascular Agent Combination Therapy” above. Additionaltherapeutic agents used to treat metabolic disorders include insulin,sulfonylureas, biguanides, alpha-glucosidase inhibitors, and incretinmimetics.

Peripheral vascular disorders are disorders related to the blood vessels(arteries and veins) located outside the heart and brain, including, forexample peripheral arterial disease (PAD), a condition that developswhen the arteries that supply blood to the internal organs, arms, andlegs become completely or partially blocked as a result ofatherosclerosis.

Gastrointestinal disorders refer to diseases and conditions associatedwith the gastrointestinal tract. Examples of gastrointestinal disordersinclude gastroesophageal reflux disease (GERD), inflammatory boweldisease (IBD), gastroenteritis, gastritis and peptic ulcer disease, andpancreatitis.

Examples of therapeutic agents used to treat gastrointestinal disordersinclude proton pump inhibitors, such as pantoprazole (Protonix),lansoprazole (Prevacid), esomeprazole (Nexium), omeprazole (Prilosec),rabeprazole; H2 blockers, such as cimetidine (Tagamet), ranitidine(Zantac), famotidine (Pepcid), nizatidine (Axid); prostaglandins, suchas misoprostoL (Cytotec); sucralfate; and antacids.

Analgesics are therapeutic agents that are used to relieve pain.Examples of analgesics include opiates and morphinomimetics, such asfentanyl and morphine; paracetamol; NSAIDs, and COX-2 inhibitors.

Pharmaceutical Compositions and Administration

Compounds provided in accordance with the present invention are usuallyadministered in the form of pharmaceutical compositions. This inventiontherefore provides pharmaceutical compositions that contain, as theactive ingredient, one or more of the compounds described, or apharmaceutically acceptable salt or ester thereof, and one or morepharmaceutically acceptable excipients, carriers, including inert soliddiluents and fillers, diluents, including sterile aqueous solution andvarious organic solvents, permeation enhancers, solubilizers andadjuvants. The pharmaceutical compositions may be administered alone orin combination with other therapeutic agents. Such compositions areprepared in a manner well known in the pharmaceutical art (see, e.g.,Remington's Pharmaceutical Sciences, Mace Publishing Co., Philadelphia,Pa. 17th Ed. (1985); and Modern Pharmaceutics, Marcel Dekker, Inc. 3rdEd. (G. S. Banker & C. T. Rhodes, Eds.)

The pharmaceutical compositions may be administered in either single ormultiple doses by any of the accepted modes of administration of agentshaving similar utilities, for example as described in those patents andpatent applications incorporated by reference, including rectal, buccal,intranasal and transdermal routes, by intra-arterial injection,intravenously, intraperitoneally, parenterally, intramuscularly,subcutaneously, orally, topically, as an inhalant, or via an impregnatedor coated device such as a stent, for example, or an artery-insertedcylindrical polymer.

One mode for administration is parenteral, particularly by injection.The forms in which the novel compositions of the present invention maybe incorporated for administration by injection include aqueous or oilsuspensions, or emulsions, with sesame oil, corn oil, cottonseed oil, orpeanut oil, as well as elixirs, mannitol, dextrose, or a sterile aqueoussolution, and similar pharmaceutical vehicles. Aqueous solutions insaline are also conventionally used for injection, but less preferred inthe context of the present invention. Ethanol, glycerol, propyleneglycol, liquid polyethylene glycol, and the like (and suitable mixturesthereof), cyclodextrin derivatives, and vegetable oils may also beemployed. The proper fluidity can be maintained, for example, by the useof a coating, such as lecithin, by the maintenance of the requiredparticle size in the case of dispersion and by the use of surfactants.The prevention of the action of microorganisms can be brought about byvarious antibacterial and antifungal agents, for example, parabens,chlorobutanol, phenol, sorbic acid, thimerosal, and the like.

Sterile injectable solutions are prepared by incorporating a compoundaccording to the present invention in the required amount in theappropriate solvent with various other ingredients as enumerated above,as required, followed by filtered sterilization. Generally, dispersionsare prepared by incorporating the various sterilized active ingredientsinto a sterile vehicle which contains the basic dispersion medium andthe required other ingredients from those enumerated above. In the caseof sterile powders for the preparation of sterile injectable solutions,the preferred methods of preparation are vacuum-drying and freeze-dryingtechniques which yield a powder of the active ingredient plus anyadditional desired ingredient from a previously sterile-filteredsolution thereof.

Oral administration is another route for administration of compounds inaccordance with the invention. Administration may be via capsule orenteric coated tablets, or the like. In making the pharmaceuticalcompositions that include at least one compound described herein, theactive ingredient is usually diluted by an excipient and/or enclosedwithin such a carrier that can be in the form of a capsule, sachet,paper or other container. When the excipient serves as a diluent, it canbe in the form of a solid, semi-solid, or liquid material (as above),which acts as a vehicle, carrier or medium for the active ingredient.Thus, the compositions can be in the form of tablets, pills, powders,lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions,syrups, aerosols (as a solid or in a liquid medium), ointmentscontaining, for example, up to 10% by weight of the active compound,soft and hard gelatin capsules, sterile injectable solutions, andsterile packaged powders.

Some examples of suitable excipients include lactose, dextrose, sucrose,sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates,tragacanth, gelatin, calcium silicate, microcrystalline cellulose,polyvinylpyrrolidone, cellulose, sterile water, syrup, and methylcellulose. The formulations can additionally include: lubricating agentssuch as talc, magnesium stearate, and mineral oil; wetting agents;emulsifying and suspending agents; preserving agents such as methyl andpropylhydroxy-benzoates; sweetening agents; and flavoring agents.

The compositions of the invention can be formulated so as to providequick, sustained or delayed release of the active ingredient afteradministration to the patient by employing procedures known in the art.Controlled release drug delivery systems for oral administration includeosmotic pump systems and dissolutional systems containing polymer-coatedreservoirs or drug-polymer matrix formulations. Examples of controlledrelease systems are given in U.S. Pat. Nos. 3,845,770; 4,326,525;4,902,514; and 5,616,345. Another formulation for use in the methods ofthe present invention employs transdermal delivery devices (“patches”).Such transdermal patches may be used to provide continuous ordiscontinuous infusion of the compounds of the present invention incontrolled amounts. The construction and use of transdermal patches forthe delivery of pharmaceutical agents is well known in the art. See,e.g., U.S. Pat. Nos. 5,023,252, 4,992,445 and 5,001,139. Such patchesmay be constructed for continuous, pulsatile, or on demand delivery ofpharmaceutical agents.

The compositions are preferably formulated in a unit dosage form. Theterm “unit dosage forms” refers to physically discrete units suitable asunitary dosages for human subjects and other mammals, each unitcontaining a predetermined quantity of active material calculated toproduce the desired therapeutic effect, in association with a suitablepharmaceutical excipient (e.g., a tablet, capsule, and ampoule). Thecompounds are generally administered in a pharmaceutically effectiveamount. Preferably, for oral administration, each dosage unit containsfrom 1 mg to 2 g of a compound described herein, and for parenteraladministration, preferably from 0.1 to 700 mg of a compound a compounddescribed herein. It will be understood, however, that the amount of thecompound actually administered usually will be determined by aphysician, in the light of the relevant circumstances, including thecondition to be treated, the chosen route of administration, the actualcompound administered and its relative activity, the age, weight, andresponse of the individual patient, the severity of the patient'ssymptoms, and the like.

For preparing solid compositions such as tablets, the principal activeingredient is mixed with a pharmaceutical excipient to form a solidpreformulation composition containing a homogeneous mixture of acompound of the present invention. When referring to thesepreformulation compositions as homogeneous, it is meant that the activeingredient is dispersed evenly throughout the composition so that thecomposition may be readily subdivided into equally effective unit dosageforms such as tablets, pills and capsules.

The tablets or pills of the present invention may be coated or otherwisecompounded to provide a dosage form affording the advantage of prolongedaction, or to protect from the acid conditions of the stomach. Forexample, the tablet or pill can comprise an inner dosage and an outerdosage component, the latter being in the form of an envelope over theformer. The two components can be separated by an enteric layer thatserves to resist disintegration in the stomach and permit the innercomponent to pass intact into the duodenum or to be delayed in release.A variety of materials can be used for such enteric layers or coatings,such materials including a number of polymeric acids and mixtures ofpolymeric acids with such materials as shellac, cetyl alcohol, andcellulose acetate.

Compositions for inhalation or insufflation include solutions andsuspensions in pharmaceutically acceptable, aqueous or organic solvents,or mixtures thereof, and powders. The liquid or solid compositions maycontain suitable pharmaceutically acceptable excipients as describedsupra. Preferably, the compositions are administered by the oral ornasal respiratory route for local or systemic effect. Compositions inpreferably pharmaceutically acceptable solvents may be nebulized by useof inert gases. Nebulized solutions may be inhaled directly from thenebulizing device or the nebulizing device may be attached to a facemasktent, or intermittent positive pressure breathing machine. Solution,suspension, or powder compositions may be administered, preferablyorally or nasally, from devices that deliver the formulation in anappropriate manner.

Synthesis of Compounds of Formula I

The compounds of the invention may be prepared using methods disclosedherein and routine modifications thereof which will be apparent giventhe disclosure herein and methods well known in the art. Conventionaland well-known synthetic methods may be used in addition to theteachings herein. The synthesis of typical compounds described herein,e.g. compounds having structures described by one or more of Formula(I), may be accomplished as described in the following Scheme A and oras provided in the following examples. If available, reagents may bepurchased commercially, e.g. from Sigma Aldrich or other chemicalsuppliers.

The 4-bromo thiazole carboxylic acid (1) is converted to thecarbohydrazide first by conversion to the acyl chloride using anacylhalide forming reagent such as thionyl chloride or oxalyl chloride.The acyl halide is then treated with hydrazine to form thecarbohydrazide (2). The carbohydrazide is cyclized in the presence ofdimethylformamide dimethylacetal and the appropriately substitutedprimary amine to afford the triazole (3) having the desired R1substituent. Alternatively, the acyl halide is converted to the amine byreaction with an appropriately substituted amine (R¹NH₂) to form thecorresponding amide of compound (1). The amide is converted to thethioamide by reaction with Lawesson's reagent using known conditions oras described herein. The thioamide is then reacted with hydrazine toafford the triazole (3). The triazole (3) is converted to the aminoanalog (intermediate A) via reaction with copper acetate and ammoniumhydroxide in DMF or other solvents known to one of ordinary skill in theart.

Preparation of the intermediate C is initiated by coupling the compound(4) with an appropriately substituted imidazole having the desired R⁶substituent. The resulting product (5) is carbonylated using palladiumreagents such as Pd (dppf)Cl₂ in the presence of carbon monoxide in aprotic solvent such as butanol. The resulting acid (6) is isolated asintermediate C. The intermediate C is then reacted with intermediate Ato form the amide intermediate D. Intermediate D is subjected to anS_(N)Ar reaction and converted to the desired amine compound of formula(I). One of ordinary skill in the art is able to perform the reactionsherein following the above general scheme, the specific proceduresprovided herein, or other literature sources known to such an artisan.

General Syntheses:

Typical embodiments of compounds in accordance with the presentinvention may be synthesized using the general reaction schemesdescribed below. It will be apparent given the description herein thatthe general schemes may be altered by substitution of the startingmaterials with other materials having similar structures to result inproducts that are correspondingly different. Descriptions of synthesesfollow to provide numerous examples of how the starting materials mayvary to provide corresponding products. Given a desired product forwhich the substituent groups are defined, the necessary startingmaterials generally may be determined by inspection. Starting materialsare typically obtained from commercial sources or synthesized usingpublished methods. For synthesizing compounds which are embodiments ofthe present invention, inspection of the structure of the compound to besynthesized will provide the identity of each substituent group. Theidentity of the final product will generally render apparent theidentity of the necessary starting materials by a simple process ofinspection, given the examples herein.

Synthetic Reaction Parameters

The terms “solvent,” “inert organic solvent” or “inert solvent” refer toa solvent inert under the conditions of the reaction being described inconjunction therewith (including, for example, benzene, toluene,acetonitrile, tetrahydrofuran (“THF”), dimethylformamide (“DMF”),chloroform, methylene chloride (or dichloromethane), diethyl ether,methanol, pyridine and the like). Unless specified to the contrary, thesolvents used in the reactions of the present invention are inertorganic solvents, and the reactions are carried out under an inert gas,preferably nitrogen.

Preparation of 2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-amine(Intermediate A)

Step 1: Preparation of 4-bromothiazole-2-carbohydrazide

To a solution of the 4-bromothiazole-2-carboxylic acid (2.0 g, 9.8 mmol)in MeOH (10 mL) was added SOCl₂ (710 μL, 9.8 mmol) and the reaction wasrefluxed for 3 hours. The reaction was concentrated, the residue wassuspended in EtOH (10 mL), hydrazine hydrate (2.4 mL, 49 mmol) was addedand the reaction was heated to reflux for 90 minutes. The reaction wasconcentrated, suspended in CH₃CN, filtered, and the solids were washedwith CH₃CN, Et₂O, and dried to afford 1.7 g (77%) of4-bromothiazole-2-carbohydrazide as a yellow solid. M+1=222.1

Step 2: Preparation of4-bromo-2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazole

4-bromothiazole-2-carbohydrazide (620 mg, 2.8 mmol) and toluene (9 mL)were added to a sealable vial, DMF-DMA (920 μL, 6.9 mmol) was added andthe reaction was stirred for 5 minutes. Cyclopropyl amine (770 μL, 11mmol), and AcOH (160 μL, 2.8 mmol) were added and the reaction washeated in a microwave reactor at 150° C. for 30 minutes. The reactionwas concentrated and purified by flash chromatography (1→7% MeOH inCH₂Cl₂) to afford 740 mg of4-bromo-2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazole (contaminatedwith DMF-DMA, but used directly in next step).

Step 3: Preparation of2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-amine

To a microwave vial was added the bromothiazole synthesized above (1.0g, 3.7 mmol), Cu(acac)₂ (97 mg, 0.37 mmol), and Cs₂CO₃ (2.4 g, 7.4 mmol)and the flask was charged with N₂. Pentadione (150 μL, 1.5 mmol), DMF (8mL), and ammonium hydroxide (1.1 ml, 300 μL/mmol) were added and thereaction was heated to 90° C. When the reaction was judged to becomplete by HPLC (˜4 hrs), the mixture was filtered through Celite, theCelite was washed with CH₂Cl₂, the filtrate was concentrated, and theresidue was purified by flash chromatography (6→13% MeOH in CH₂Cl₂) toprovide 480 mg (63% over two steps) of2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-amine as an oil.M+1=208.2

Preparation of 2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-amine(Intermediate B)

Step 1: Preparation of 4-bromothiazole-2-carboxylic Acid

To the solution of 2,4-dibromothiazole (50 g, 207 mmol, 1.0 eq.) in Et20(1000 ml) was added n-BuLi (90 ml, 2.5 M, 1.1 eq.) at −78° C. dropwiseand it was stirred for one hour. The reaction solution was poured intodry CO₂ at −78° C. and the reaction mixture was warmed to the roomtemperature. TLC and LCMS showed the reaction was complete. It wasquenched with water (100 ml). The Et20 phase was removed. The aqueousphase was adjusted to pH to 2-3 and extracted with ethyl acetate. Theorganic phase was dried, filtered and concentrated to obtain 35 g (82%y) of the 4-bromothiazole-2-carboxylic acid. ¹HNMR (400 MHz, DMSO):δ8.23 (1H, s)

Steps 2-3: Preparation of(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carboxamide

To the mixture of 4-bromothiazole-2-carboxylic acid (80 g, 1.0 eq.) inSOCl₂ (300 ml) was added DMF (10 drops). It was refluxed for 5 hours andconcentrated. The residue was dissolved in DCM (300 ml) and added to thesolution of (S)-1,1,1-trifluoropropan-2-amine hydrochloride (60.5 g,1.05 eq.) with Et3N (117 g, 3.0 eq.) at 0° C. It was stirred overnight.TLC and LCMS showed the reaction was complete. It was quenched withwater and extracted with DCM. The organic phase was dried, filtered andconcentrated. The residue was purified by column to obtain 62 g (53% y)of(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carb oxamide.

1HNMR (400 MHz, CDCl3): δ7.47 (1H, s), 7.17 (1H, br s), 4.74-4.76 (1H,m), 1.39 (3H, d, J=6.8 Hz).

Step 4: Preparation of(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carbothioamide

To a solution of(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carboxamide (50 g,1.0 eq.) in toluene (1000 ml) was added Lawesson's Reagent (100 g, 1.5eq.). It was refluxed overnight. TLC and LCMS showed the reaction wascomplete. It was concentrated and the residue was purified by column toobtain 46 g (88% y) of(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carbothioamide.

Step 5: Preparation of(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carbohydrazonamide

To a solution of(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carbothioamide (46g, 1.0 eq.) was added hydrazine hydrate (15 g, 2.0 eq.). It was refluxedovernight. TLC and LCMS showed the reaction was complete. It wasconcentrated and the residue was purified by a flash column to obtain 48g (100% y) of(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carbohydrazonamide

Step 6: Preparation of(S)-4-bromo-2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazole

(S)-4-bromo-N-(1,1,1-trifluoropropan-2-yl)thiazole-2-carbohydrazonamide(55 g,) in triethoxymethane (500 ml) was stirred at 90° C. for 3 hoursand then at 130° C. overnight and the reaction was concentrated. Theresidue was purified by column to obtain 44 g (77% y) of(S)-4-bromo-2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazole.

¹HNMR (400 MHz, CDCl3): δ8.46 (1H, s), 7.43 (1H, s), 6.48-6.52 (1H, m),1.83 (3H, d, J=7.2 Hz).

Step 7: Preparation of(S)-2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-amine(Intermediate B)

To a solution of(S)-4-bromo-2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazole(50 g, 120 mmol, 1.0 eq.) in DMF (500 ml) was added Cu(acac)₂ (3.2 g, 12mmol, 0.1 eq.), acetylacetone (1.2 g, 0.1 eq.) and NH₄OH (50 ml, conc).It was stirred at 90° C. overnight and it was concentrated. The residuewas dissolved in MeOH (500 ml) and it was filtered. The filtrate wasconcentrated and the residue was purified by column to obtain 9.6 g (24%y) of Intermediate B.

¹HNMR (400 MHz, CDCl3): δ8.38 (1H, s), 6.58-6.65 (1H, m), 6.14 (1H, s),4.20 (2H, br s), 1.77 (3H, d, J=7.2 Hz); ESI MS: 264 ([M+1]).

Preparation of 4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropicolinic Acid(Intermediate C)

Step 1: Preparation of2-chloro-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropyridine

A suspension 2-chloro-5-fluoro-4-iodopyridine (1.8 g, 1.00 mmol),4-cyclopropyl imidazole (982 mg, 9.10 mmol), Cu₂O (100 mg, 0.700 mmol),8-hydroxyquinoline (152 mg, 1.05 mmol), cesium carbonate (4.60 g, 14.0mmol), and PEG-3350 (1.4 g) in butyronitrile (50 mL) was heated at 65°C. for 16 hours. The reaction mixture was filtered through Celite,concentrated and the residue was partitioned between dichlormethane andwater. The layers were separated and the aqueous layer washed twice withdichloromethane. The combined organic layers were dried (MgSO₄),filtered and concentrated. The residue was purified by flashchromatography (15→60% EtOAc in hexanes) to afford2-chloro-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropyridine (702 mg,42% yield).

Step 2: Preparation of butyl4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropicolinate

2-chloro-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropyridine (730 mg,3.07 mmol) and Pd(dppf)Cl₂ (112 mg, 0.159 mmol) were suspended indegassed BuOH, and the reaction vessel was purged with carbon monoxideand a balloon of CO was affixed to the reaction vessel. The mixture washeated to 70° C. for 90 minutes, filtered through Celite, andconcentrated. The residue was passed through a short plug of silica gelto afford butyl 4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropicolinate(860 mg, 93%).

Step 3: Preparation of4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropicolinic Acid

4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropicolinate (640 mg, 2.11mmol) was dissolved in 1N HCl (5 mL) and the reaction was heated to 100°C. overnight. The solvent was removed, CH₃CN was added and the solventremoved to afford 4-(4-cyclopropyl-1H-imidazol-1-yl)-5-fluoropicolinicacid which was used directly in subsequent reactions (assuming 100%conversion).

Representative Procedure for Amide Coupling Reaction Preparation of4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-fluoropicolinamide(Intermediate D)

To a mixture of intermediate C.HCl (340 mg, 1.2 mmol), intermediate A(261 mg, 1.26 mmol), HATU (638 mg, 1.68 mmol), and N-methyl morpholine(330 μL, 3.00 mmol) was added DMF (5 mL) and the reaction was stirredfor 2 hours. The mixture was concentrated, redissolved in a minimalamount of CH₃CN, and water was added dropwise until a thick slurry wasformed. The solids were isolated by filtration and washed with CH₃CN toafford 320 mg (61% y). M+1=437.2

Intermediate E was Prepared According to the Same Procedure

Representative Procedure for S_(N)Ar Displacement of Aryl Fluoride toYield Final Products Preparation of4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-morpholinopicolinamide

To a mixture of intermediate D (450 mg, 1.03 mmol) and K₂CO₃ (430 mg,3.09 mmol) in DMF (3 mL) was added morpholine (270 Ld, 3.09 mmol) andthe reaction was heated to 80° C. overnight. The reaction wasconcentrated and purified by RP-HPLC to afford 390 mg (75% y) of4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-morpholinopicolinamide.All final compounds were isolated as the HCl salt from the preparativeHPLC.

C₂₄H₂₅N₉₀O₂S. 504.2. (M+1). ¹H NMR (DMSO) δ 10.99 (s, 1H), 9.57 (s, 1H),8.79 (s, 1H), 8.64 (s, 1H), 8.27 (s, 1H), 7.99 (s, 2H), 4.05-4.09 (m,1H), 3.61-3.64 (m, 4H), 2.90-3.05 (m, 4H), 2.04-2.08 (m, 1H), 1.03-1.09(m, 6H), 0.85-0.90 (m, 2H).

The Following Compounds were Prepared in a Similar Fashion Using theAppropriate Amine and the Indicated Intermediate

Prepared Via Intermediate D

4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-(3-hydroxy-3-methylazetidin-1-yl)picolinamideC₂₄H₂₅N₉O₂S. 504.2. (M+1). ¹H NMR (DMSO) δ 10.74 (s, 1H), 9.32 (s, 1H),8.76 (s, 1H), 8.14 (s, 1H), 8.10 (s, 1H), 7.93 (s, 1H), 7.77 (s, 1H),4.02-4.06 (m, 1H), 3.67 (dd, J=8.0, 20.4 Hz, 4H), 2.01-2.06 (m, 1H),1.37 (s, 3H), 1.03-1.10 (m, 6H), 0.83-0.86 (m, 2H).

Prepared Via Intermediate E

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-morpholino-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamideC₂₄H₂₄F₃N₉O₂S. 560.2. (M+1). ¹H NMR (DMSO) δ 11.46 (s, 1H), 9.56 (s,1H), 9.23 (s, 1H), 8.64 (s, 1H), 8.27 (s, 1H), 8.01 (s, 1H), 7.99 (s,1H), 6.78-6.82 (m, 1H), 3.61-3.65 (m, 4H), 2.90-2.94 (m, 4H), 2.02-2.06(m, 1H), 1.83 (d, J=7.2 Hz, 3H), 1.03-1.07 (m, 2H), 0.86-0.88 (m, 2H).

Prepared Via Intermediate E

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(3-hydroxy-3-methylazetidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamideC₂₄H₂₄F₃N₉O₂S. 560.1. (M+1). ¹H NMR (DMSO) δ 11.21 (s, 1H), 9.24 (br s,1H), 9.22 (s, 1H), 8.13 (s, 1H), 8.09 (s, 1H), 7.96 (s, 1H), 7.75 (s,1H), 6.79 (pent, J=7.2 Hz, 1H), 3.67 (dd, J=8.0, 19.6 Hz, 4H), 2.68 (s,1H), 2.02-2.05 (m, 1H), 1.82 (d, J=7.2 Hz, 3H), 1.37 (s, 3H), 1.01-1.06(m, 2H), 0.81-0.85 (m, 2H).

Prepared Via Intermediate E

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(dimethylamino)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamideC₂₂H₂₂F₃N₉OS. 518.1. (M+1). ¹H NMR (DMSO) δ 11.32 (s, 1H), 9.43 (s, 1H),9.23 (s, 1H), 8.55 (s, 1H), 8.17 (s, 1H), 7.98 (s, 1H), 7.84 (s, 1H),6.80 (pent, J=7.2 Hz, 1H), 2.73 (s, 6H), 2.02-2.06 (m, 1H), 1.83 (d,J=7.2 Hz, 3H), 1.01-1.06 (m, 2H), 0.84-0.88 (m, 2H).

Additional General Preparation of Compounds of Formula (I)

wherein R^(a), R^(b), and R¹ are defined in formula (I).

In a 2 mL reaction vial was placed the appropriate amine nucleophile(4.1 equivalents) and K₂CO₃ (6.1 equivalents). To this was added asolution of Intermediate D or E (1.0 equivalent) in NMP (0.15 M). Themixture was heated at 80° C. for 16 h. It was cooled to roomtemperature, and to this was added EtOAc. The resulting supernatant wastransferred to a separate collection tube. The solvent was removed byGenevac, and the resulting solid was washed with water and dried to givethe desired product. The product identity was verified by LCMS analysis.

Preparation of4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(3-hydroxypyrrolidin-1-yl)pyridine-2-carboxamide

In a 2 ml reaction vial was placed pyrrolidin-3-olhydrochloride (22.7mg, 0.19 mmol) and K₂CO₃ (38 mg, 0.28 mmol). To this was added asolution of Intermediate D (20 mg, 0.046 mmol) in NMP (0.3 mL). Themixture was heated at 80° C. for 16 h. It was cooled to roomtemperature, and to this was added EtOAc (1 mL). The solvent was removedby Genevac, and the resulting solid was washed with water and dried togive4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(3-hydroxypyrrolidin-1-yl)pyridine-2-carboxamide(Compound 10). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₄H₂₆N₉O₂S, 504.2,504.1.

Compounds 6, 9, 12, 15 and 17 were prepared in a similar fashion usingIntermediate D.

5-(azetidin-1-yl)-4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]pyridine-2-carboxamide(Compound 6). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₃H₂₃N₉OS 474.2, found:474.2.

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-pyrrolidin-1-ylpyridine-2-carboxamide(Compound 9). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₄H₂₅N₉OS 488.2, found:488.2.

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-piperidin-1-ylpyridine-2-carboxamide(Compound 12). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₅H₂₇N₉OS 502.2, found:502.1.

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(4-hydroxy-4-methylpiperidin-1-yl)pyridine-2-carboxamide(Compound 15). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₆H₂₉N₉O₂S, 532.2,found: 532.1.

4-(4-cyclopropylimidazol-1-yl)-N-[2-(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(4-methylpiperazin-1-yl)pyridine-2-carboxamide(Compound 17). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₅H₂₈N₁₀OS 517.2,found: 517.1.

Compounds 7, 8, 11, 13, 14, and 16 were prepared in similar generalpreparation procedure using Intermediate E.

(S)-5-(azetidin-1-yl)-4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 7). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₃H₂₂F₃N₉OS 530.2,found: 530.2.

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(pyrrolidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 8). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₄H₂₄F₃N₉OS 544.2,found: 543.8.

4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(3-hydroxypyrrolidin-1-yl)-N-(2-(4-((S)-1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 11). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₄H₂₄F₃N₉O₂S, 560.2,found: 560.2.

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(piperidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 13). LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₅H₂₆F₃N₉OS 558.2,found: 557.9.

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(4-hydroxy-4-methylpiperidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 14). LRMS (ESI⁺) m/z [M+H] calcd for C₂₆H₂₈F₃N₉O₂S, 588.2,found: 587.9.

(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(4-methylpiperazin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide(Compound 16) LRMS (ESI⁺) m/z [M+H]⁺ calcd for C₂₅H₂₇F₃N₁₀OS 573.2,found: 573.1.

Utility, Testing and Administration

General Utility

The compounds of Formula I are believed effective in the treatment ofconditions that respond to administration of ASK1 inhibitors.Specifically, the compounds of Formula I are useful in the treatment ofa broad range of diseases, for example autoimmune disorders,inflammatory diseases, cardiovascular diseases (including diabetes,diabetic nephropathy, and other complications of diabetes), cardio-renaldiseases, including kidney disease, fibrotic diseases, respiratorydiseases (including COPD, idiopathic pulmonary fibrosis (IPF), and acutelung injury), acute and chronic liver diseases, and neurodegenerativediseases.

Testing

Activity testing is conducted as described in the Examples below, and bymethods apparent to one skilled in the art.

The following examples are included to demonstrate preferred embodimentsof the invention. It will be appreciated by those of skill in the artthat the techniques disclosed in the examples which follow representtechniques discovered by the inventor(s) to function well in thepractice of the invention, and thus can be considered to constitutepreferred modes for its practice. However, those of skill in the artshould, in light of the present disclosure, appreciate that many changescan be made in the specific embodiments which are disclosed and stillobtain a like or similar result without departing from the spirit andscope of the invention.

Biological Assays

To evaluate the inhibitory activity of compounds of the inventionagainst ASK1 (Apoptosis Signal-regulating Kinase 1) kinase, its activitywas examined using a TR-FRET ASK1 kinase assay which determined theamount of phosphate transferred to a peptide substrate from ATP.

Materials and Methods

Reagents

Dephosphorylated recombinant human ASK1 kinase was from Gilead Sciences.Small molecule kinase inhibitor staurosporine (Catalogue # S6942) anddithiothreitol (DTT, catalogue #43815-5G) were obtained from SigmaChemicals (St. Louis, Mo.). ATP (catalogue #7724) was from Affymetrix(Santa Clara, Calif.) and test compounds were from Gilead Sciences. HTRFKinEASE™-STK S3 kit was obtained from Cisbio (Bedford, Mass.). All otherreagents were of the highest grade commercially available.

Assays

The assay measures the phosphorylation level of a biotinylated peptidesubstrate by the ASK1 kinase using HTRF detection (6.1). This is acompetitive, time-resolved fluorescence resonance energy transfer(TR-FRET) immunoassay, based on HTRF® KinEASE™-STK manual from Cisbio(6.1). Test compound, 1 μM STK3 peptide substrate, 4 nM of ASK1 kinaseare incubated with 10 mM MOP buffer, pH. 7.0 containing 10 mMMg-acetate, 0.025% NP-40, 1 mM DTT, 0.05% BSA and 1.5% glycerol for 30minutes then 100 μM ATP is added to start the kinase reaction andincubated for 3 hr. Peptide antibody labeled with 1×Eu³⁺ Cryptate buffercontaining 10 mM EDTA and 125 nM Streptavidin XL665 are added to stopthe reaction and phosphorylated peptide substrate is detected usingEnvision 2103 Multilabeled reader from PerkinElmer. The fluorescence ismeasured at 615 nm (Cryptate) and 665 nm (XL665) and a ratio of 665nm/615 nm is calculated for each well. The resulting TR-FRET level (aratio of 665 nm/615 nm) is proportional to the phosphorylation level.Under these assay conditions, the degree of phosphorylation of peptidesubstrate was linear with time and concentration for the enzyme. Theassay system yielded consistent results with regard to K_(m) andspecific activities for the enzyme. For inhibition experiments (IC₅₀values), activities were performed with constant concentrations of ATP,peptide and several fixed concentrations of inhibitors. Staurosporine,the nonselective kinase inhibitor, was used as the positive control. Allenzyme activity data are reported as an average of quadruplicatedetermination.

Data Analysis

The IC₅₀ values were calculated following equation:y=Range/{1+(x/IC ₅₀)^(s)}+BackgroundWhere x and y represent the concentration of inhibitors and enzymeactivity, respectively. Enzyme activity is expressed as the amount ofPhosphate incorporated into substrate peptide from ATP. Range is themaximum y range (no inhibitor, DMSO control) and s is a slope factor(6.2).Results

Following the above method, tested compounds of Formula (I) inhibitedASK1 as shown below.

ASK1 Example # Compound name IC₅₀ (nM) 14-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4- 1.44cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-morpholinopicolinamide 24-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4- 1.58cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-(3-hydroxy-3-methylazetidin-1- yl)picolinamide 3(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5- 2.62morpholino-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4- yl)picolinamide 4(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(3- 1.69hydroxy-3-methylazetidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide 5(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5- 1.88(dimethylamino)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3- yl)thiazol-4-yl)picolinamideStaurosporine 6.30

The data demonstrates that tested compounds of formula (I) are potentinhibitors of the ASK-1 receptor.

ASK1 (Apoptosis Signal-Regulating Kinase 1) 293 Cell-Based Assay(Cellular EC₅₀)

The cellular potency of compounds was assayed in cells stably expressingan AP-1:luciferase reporter construct (293/AP1-Luc cells—Panomics Inc.,6519 Dumbarton Circle, Fremont, Calif.). Cells were infected with anadenovirus expressing kinase active ASK1 (631-1381 of rat ASK1 cDNA),which will activate the AP-1 transcription factor and increase theexpression of luciferase. Inhibitors of ASK1 will decrease the enzymeactivity of ASK1 and therefore decrease the activity of AP-1transcription factor and the expression of luciferase.

1. Materials Required for this Protocol

Media and Reagents Source Company Catalog No. AP-1 Reporter 293 StableCell Panomics Unknown Line DMEM (w/ high glucose, w/o L- MediaTech15-018-CM glutamine, w/ pyruvate, w/ HEPES DMEM (w/ high glucose, w/o L-Invitrogen 31053-028 glutamine, w/o pyruvate, w/o HEPES, w/o phenol redHEPES, 1M Invitrogen 15630-080 Sodium Pyruvate, 100 mM Invitrogen11360-070 Fetal Bovine Serum, “FBS” Hyclone SH30088.03 Pen-Strep-Glut.,“PSG” Invitrogen 10378-016 HygromycinB Calbiochem 400052 Dulbecco's PBS(sterile) MediaTech 21-030-CM Trypsin-EDTA (0.25%) Invitrogen 25200-056Steady-Glo Luciferase Assay Promega E2550 System Labware Source CatalogNo. Flasks (poly-D-Lysine coated, 150 BD Biosciences 356538 cm², ventedcap) Plates (poly-D-Lysine coated, Greiner (through 781944 384-well,white/clear, sterile TCT) VWR Scientific) (82051-354) White Backing TapePerkinElmer 6005199 Cell Strainers (40 um nylon, blue VWR Scientific21008-949 ring, fits 50 mL conical vials)2. Reference Materials1. Panomics 293/AP1-Luc stable cell-line product insert.2. Promega Steady-Glo Luciferase Assay System product insert.3. Media Required3. Complete Growth Medium, “CGM”a. DMEM (MediaTech)b. 10% FBSc. 1% PSGd. 100 ug/mL HygromycinB4. Assay Medium, “AM”a. DMEM (Invitrogen)b. 25 mM HEPESc. 1 mM Sodium Pyruvated. 1% PSG4. MethodsMaintenance:1. 293/AP1-Luc Maintain 293/Acells per vendor's instructions; harvestcells at ˜80% confluence in T150 flasks as follows:a. Aspirate media, wash gently with ˜12 mL sterile D-PBS, aspirate.b. Add 5 mL Trypsin-EDTA, tilt gently to coat flask, and incubate ˜5min. at 37° C.c. Do not tap flask; add 5 mL CGM, wash flask 4× with cell suspension,transfer to 50 mL conical vial, centrifuge 5 min. at 1200 rpm.d. Aspirate media from cell pellet, add 20 to 30 mL CGM, resuspendpellet by pipetting 6×, pass through cell strainer to disperse clumps(if necessary), and count cells with hemocytometer.Assay Day 1:2. Harvest cells as above, except resuspend cell pellet.Count cells and dilute to 1.5×10⁵ cells per mL; add adenovirus such thatthere are 5 infectious forming units per cell.3. Prime (20 to 30 mL) and plate cells in Greiner poly-D-Lysine coated384-well plates at 1.2×10⁴ cells per well using BioTek uFill (80 uL perwell).4. Immediately dose plates with 0.4 uL of compound dose series (in 100%DMSO) incubate 24 hours in humidified incubator (37° C., 5% CO₂).Assay Day 2:5. Process plates (per manufacturer's instructions) as follows:a. Set plates in laminar flow hood & uncover for 30 minutes at roomtemperature to cool.b. Remove 60 uL of AM from assay wellsc. Add 20 uL per well Steady-Glo Firefly substrate, let sit for 10-20minutes at room temperatured. Cover bottom of assay plates with white backing tape.e. Acquire data on a fluorescence plate reader

The 100% inhibition positive control wells were generated by infectingcells with an adenovirus expressing catalytically inactive ASK1 mutantwith lysine to argine mutation at residue 709.

Result

ASK1 Example EC50 # Compound name (nM) 14-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4- 13.15cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-morpholinopicolinamide 2 4-(4-cyclopropyl-1H-imidazol-1-yl)-N-(2-(4-11.17 cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)-5-(3-hydroxy-3-methylazetidin-1- yl)picolinamide 3(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5- 6.74morpholino-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4- yl)picolinamide 4(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5-(3- 5.21hydroxy-3-methylazetidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3- yl)thiazol-4-yl)picolinamide 5(S)-4-(4-cyclopropyl-1H-imidazol-1-yl)-5- 7.12(dimethylamino)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3- yl)thiazol-4-yl)picolinamide

The above data suggests that compounds of formula (I), particularlycompounds tested exhibit potent in-vitro efficacy.

Human Whole Blood CXCL1 Assay

Reagents

Human blood collected in sodium heparin glass vacutainer tubes (BDBiosciences #366480) was obtained from Stanford Blood Center (Palo Alto,Ca). Red blood cell Ammonium-Chloride-Potassium (ACK) lysis buffer wasobtained from Invitrogen (A10492-01) Phosphate buffered saline (PBS,21-031-CV) and RPMI640 media (10-040-CV) were purchased from Cellgro. 50ml conical tubes were from BD Biosciences (352070). Multiscreen Filterplates (MSBVS 1210) used along with vacuum manifold for 96-well plate(MAVM0960R) were purchased from EMD Millipore Inc. Transfer pipettes(13-711-20) and hemocytometer (0267110) were purchased from ThermoFisher Scientific. An Allegra X15R from Beckmann Coulter was used forall the centrifugation steps in the isolation of PBMCs.

ASK1 inhibitors and Auranofin (Enzo Life Sciences Inc. BML-EI206-0100)were dissolved in dimethyl sulfoxide (DMSO, Sigma Aldrich cat#472301) ata concentration of 10 mM and 100 mM, respectively. Aliquots were storedfrozen at −20° C. until the time of use and not reused or refrozen.Serial dilutions of ASK1 inhibitors were preformed in saline (pH 4.5-7,Hospira Inc, RL-2099). Cell Extraction buffer (FNN0011) and Haltprotease/phosphatase inhibitor cocktail (78442) were purchased from LifeTechnologies, Inc. and Thermofisher Scientific Inc., respectively. ABoekel scientific Rocker II was used to agitate the plates betweentreatments. CXCL1 protein was quantified with an ELISA assay fromMillipore.

Compound Treatment and Auranofin Stimulation

A 10 mM ASK1 inhibitor DMSO stock was diluted in saline to create a 100μM (10×) working solution. This was subjected to an eight point,three-fold serial dilution series in saline, with each concentration at10× the final intended concentration. 10 uL of each 10× working stockwas added to eight wells of a 96 plate containing 90 μl of whole blood,creating a dose range of 10 uM to 1.5 nM (Final assay plateconfiguration shown below) with eight replicates per concentration. Thesamples were incubated for one hour in a 37° C. and 5% CO₂ incubator. A100 mM Auranofin stock was diluted to 100 μM in RPMI media and 5 ul wasadded to each well of the assay plate to make a final concentration ofapproximately 5 μM. In control samples that were not treated with Ask1inhibitor or Auranofin, DMSO/saline were added at equivalent amounts asthe wells incubated with the highest concentration of test compound(0.1% final concentration of DMSO). The samples were incubated in a 37°C./5% CO₂ incubator for 24 hours with gentle agitation. At the end ofthe incubation period, 100 μl of RPMI 1640 media was added to each well,and the plates were gently mixed and then centrifuged at 1000×g for 10min at 4° C. to pellet the cells. 50 μl of the resulting supernatantfrom each well was transferred to a new 96 well assay plate and theamount of CXCL1 was quantified by a Procarta immunoassay following themanufacturer's instructions. Each compound was tested in blood from atleast two independent donors.

Final Assay Plate Configuration

Test Compound μM 0 0 0.0045 0.014 0.041 0.12 0.37 1.1 3.3 10 AuranofinμM 0 5 5 5 5 5 5 5 5 5 Replicate 1 Replicate 2 Replicate 3 Replicate 4Replicate 5 Replicate 6 Replicate 7 Replicate 8Data Analysis

Data was analyzed using GraphPad Prism 5 software (GraphPad SoftwareInc., La Jolla, Calif., USA) using nonlinear regression curve fittingwith variable slope to determine the EC₅₀ values. The average EC₅₀ fromat least two independent donors is reported.

Results:

Example WB EC50 nM No. Compound Name (n = 4) 14-(4-cyclopropyl-1H-imidazol- 243 1-yl)-N-(2-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)thiazol-4-yl)- 5-morpholinopicolinamide 3(S)-4-(4-cyclopropyl-1H- 202 imidazol-1-yl)-5-morpholino-N-(2-(4-(1,1,1-trifluoropropan- 2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide

The data suggests that compounds of formula (I) are potent in vivoinhibitors of ASK1. The activities of Compounds 6-17 were examined usingthe assay measures the phosphorylation level of a biotinylated peptidesubstrate by the ASK1 kinase using the same competitive TR-FRETimmunoassay based on HTRF® KinEASE™-STK manual from Cisbio (6.1) asdescribed above. The tested compounds inhibited ASK1 as shown below.

ASK1 Example # Compound name IC₅₀ (nM) 6 5-(azetidin-1-yl)-4-(4- 2.4cyclopropylimidazol-1-yl)-N-[2-(4- cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]pyridine-2-carboxamide 7(S)-5-(azetidin-1-yl)-4-(4-cyclopropyl- 1.71H-imidazol-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol- 3-yl)thiazol-4-yl)picolinamide 8(S)-4-(4-cyclopropyl-1H-imidazol-1- 2.2yl)-5-(pyrrolidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol- 3-yl)thiazol-4-yl)picolinamide 94-(4-cyclopropylimidazol-1-yl)-N-[2- 1.9(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-pyrrolidin-1-ylpyridine- 2-carboxamide 104-(4-cyclopropylimidazol-1-yl)-N-[2- 3(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(3-hydroxypyrrolidin-1- yl)pyridine-2-carboxamide 114-(4-cyclopropylimidazol-1-yl)-5-(3- 2.2hydroxypyrrolidin-1-yl)-N-[2-[4- (1,1,1-trifluoropropan-2-yl)-1,2,4-triazol-3-yl]-1,3-thiazol-4-yl]pyridine- 2-carboxamide 124-(4-cyclopropylimidazol-1-yl)-N-[2- 2(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-piperidin-1-ylpyridine- 2-carboxamide 13(S)-4-(4-cyclopropyl-1H-imidazol-1- 0.7yl)-5-(piperidin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol- 3-yl)thiazol-4-yl)picolinamide14 4-(4-cyclopropylimidazol-1-yl)-5-(4- 1.8hydroxy-4-methylpiperidin-1-yl)-N-[2-[4-(1,1,1-trifluoropropan-2-yl)-1,2,4-triazol-3-yl]-1,3-thiazol-4-yl]pyridine- 2-carboxamide 154-(4-cyclopropylimidazol-1-yl)-N-[2- 2.5(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3- thiazol-4-yl]-5-(4-hydroxy-4-methylpiperidin-1-yl)pyridine-2- carboxamide 16(S)-4-(4-cyclopropyl-1H-imidazol-1- 2.1yl)-5-(4-methylpiperazin-1-yl)-N-(2-(4-(1,1,1-trifluoropropan-2-yl)-4H-1,2,4-triazol-3-yl)thiazol-4-yl)picolinamide 174-(4-cyclopropylimidazol-1-yl)-N-[2- 3.1(4-cyclopropyl-1,2,4-triazol-3-yl)-1,3-thiazol-4-yl]-5-(4-methylpiperazin-1- yl)pyridine-2-carboxamide

The data demonstrates that tested compounds of formula (I) are potentinhibitors of the ASK-1 receptor.

What is claimed is:
 1. A compound having the structure: